The aim of the project is to investigate the role of intracellular-free calcium in the response to cells to hyperthermia. Experiments are designed to determine the effect of clinically relevant heating doses on cell Ca2+ concentrations and to elucidate the subcellular origins of heat-induced Ca2+ increase. Studies will further trace the mechanisms involved in the mobilization of Ca2+ from internal pools. Effects on membrane phosphoinositides and on mitochondrial and endoplasmic reticulum Ca2+ gating will be explored. Events distal to cell Ca2+ increases including activation of protein kinases, proteases and phospholipases will be investigated in an effort to gain an understanding of the role of Ca2+ in pathways of thermal cell killing. Changes in ion concentration and enzyme activities will be correlated with parallel studies on clonogenicity. The ultimate purpose of the study is to gain a clearer understanding of the mechanism of hyperthermic killing. This may permit a more rational application of hyperthermia alone and in combination with other treatments. In addition, a role for Ca2+ some classes of active drugs which are not currently used for cancer treatment will be tested in the study.